With over 2000 deaths each year in the United States, Sudden Infant Death Syndrome (SIDS) remains the leading cause of death in infants 1 month to 1 year of age. The fundamental causes of SIDS remain poorly understood. Current etiologic studies for SIDS have focused on the central/autonomic nervous system, inborn errors of metabolism, and genetic cardiac channelopathies as possible substrates for the "vulnerable infant" in accordance with the SIDS triple risk hypothesis. Our prior work investigating Cardiac Channel Mutations in SIDS (HD42569) has established that the pathogenic mechanism for approximately 10% of SIDS may stem from the most common cardiac channelopathy known as long QT syndrome (LOTS). Recently, our Sudden Death Genomics Laboratory has discovered and functionally characterized two novel LQTS-susceptibility genes: CAV3 (LQT9) and SCN4B (LQT10) involving mutations in Channel Interacting Proteins (ChlPs) of the cardiac sodium channel macromolecular complex yielding LQTS-phenotypes. Most recently, mutation scanning of a limited region of the encoded calcium release channel revealed pathogenic mutations in approximately 2% of white infants. Functional studies indicate an arrhythmogenic mechanism consistent with type 1 catecholaminergic polymorphic ventricular tachycardia (CPVT1). In addition, polymorphisms that might predispose infants to sudden death in critical situations have been identified in various signaling pathways of the autonomic nervous system (ANS). However, these initial observations, derived from small cohorts of SIDS, have not been confirmed. Given the relationship between prolongation of the QT interval and dysregulation of the ANS, this proposal will seek to confirm the associations between SIDS and genetic variation in critical components of the ANS. These discoveries frame the experiments outlined in this renewal and support the applicant's hypothesis that an additional 5 - 10% of SIDS may stem from mutations in the ChlPs that comprise the sodium channel macromolecular complex and the calcium release channel macromolecular complex. If correct, this proposal will provide molecular and functional evidence implicating mutations in either the cardiac channel pore-forming subunits or ChlPs as the probable cause for 20% of SIDS (an estimated 400 cases each year). Furthermore, it will explore the involvement of genetic variations in genes encoding key proteins of the ANS- related serotonergic signaling pathways in the pathogenesis of this disease. Specifically, using polymerase chain reaction, denaturing high performance liquid chromatography, and DNA sequencing on one of the world's largest DNA cohorts of SIDS cases (N = 600), the applicant proposes to i) conduct comprehensive open reading frame/splice site mutational analysis of 12 genes that encode key ChlPs within the sodium channel and calcium release channel macromolecular complexes as candidate genes for SIDS and ii) conduct confirmatory case-control association studies involving polymorphisms/genetic variations in the ANS as well as novel candidate gene mutation analysis of VMAT2, a gene that encodes a key determinant of ANS-related serotonergic signaling.